The cellular and molecular mechanisms of free radical-related pancreatic islet destruction are incompletely understood. Pharmacological modulation of this islet cell death may provide novel avenues for the experimental therapy of diabetes. Recent studies in isolated islet cells, and other cell types demonstrate that peroxynitrite and hydrogen peroxide, potent oxidants produced during immune-cell mediated islet cell attack, trigger an intracellular cascade culminating in cellular energy failure. DNA strand-breakage triggered by hydrogen peroxide and peroxynitrite is recognized by a specific domain of the nuclear enzyme poly (ADP-ribose) synthetase (PARS). This results in the activation of PARS which initiates an energy consuming inefficient repair cycle, with resultant depletion of dinucleotide pools, slowing the rate of glycolysis and mitochondrial respiration, reducing ATP synthesis. Thus, PARS acts as a terminal mediator of cellular energetic collapse leading to cellular necrosis. The preliminary data show that pharmacological inhibition of PARS with 5-iodo-6-amino-1,2 benzopyrone (INH2BP), a novel, potent inhibitor of PARS reduces the development of hyperglycemia and islet cell destruction in a rot model of streptozotocin-induced diabetes. Furthermore, data from the group, and also from two other groups of investigators demonstrate that PARS deficient (knockout) mice are resistant against streptozotocin-induced diabetes. Because the streptozotocin-model is not fully predictable of the human situation, the first aim of the current protocol is aimed at investigating the role of PARS activation in the pathophysiology of spontaneous autoimmune diabetes: PARS deficient mice will be back-crossed with the NOD mice, and the development of diabetes will be investigated in this model. The course of pancreatic insulin depletion, the course of insulitis, and the development of hyperglycemia will be monitored. They will also test the effect of the PARS inhibitor INH2BP on the course of diabetes development in the NOD model. The second aim of the proposed studies is to perform in vitro studies in islets from wild type and PARS deficient mice in order to define the mode of cell death (necrosis and apoptosis) induced by peroxynitrite, and to define the role of PARS in the process. The current proposal will provide (1) novel mechanistic information on the mechanism of autoimmune diabetes, and (2) will result in data which will help the company in making a strategic decision for the preclinical development of a novel, potent, orally active PARS inhibitor.